AU2012201208B2 - Microbicidal composition - Google Patents

Abstract

Synergistic microbicidal compositions containing N-(n-butyl)-1,2 benzisothiazolin- 3-one.

Description

AUSTRALIA Patents Act COMPLETE SPECIFICATION (ORIGINAL) Class Int. Class Application Number: Lodged: Complete Specification Lodged: Accepted: Published: Priority Related Art: Name of Applicant: Rohm and Haas Company Actual Inventor(s): Megan Anne Diehl, Richard Levy, Dolores Ann Shaw, Eileen Fleck Warwick Address for Service and Correspondence: PHILLIPS ORMONDE FITZPATRICK Patent and Trade Mark Attorneys 367 Collins Street Melbourne 3000 AUSTRALIA Invention Title: MICROBICIDAL COMPOSITION Our Ref: 931035 POF Code: 496570/83886 The following statement is a full description of this invention, including the best method of performing it known to applicant(s): -1 Ocoeq -1a MICROBICIDAL COMPOSITION The present application is a divisional application from Australian Patent Application Number 2006222721, which claims priority from European Patent 5 Application Number 05292073.3, filed 4 October 2005, the entire disclosure of which is incorporated herein by reference. This invention relates to a synergistic combination of selected microbicides having greater activity than would be observed for the individual microbicides. In some cases, commercial microbicides cannot provide effective control of 0 microorganisms, even at high use concentrations, due to weak activity against certain types of microorganisms, e.g., those resistant to some microbicides, or due to aggressive environmental conditions. Combinations of different microbicides are sometimes used to provide overall control of microorganisms in a particular end use environment. For example, U.S. Pat. App. Pub. No. 5 2004/0014799 discloses a synergistic combination of N-(n-butyl)-1,2 benzisothiazolin-3 -one (BBIT) and 2 -methyl-4-isothiazolin- 3-one (MI) over the limited range of ratios of BBIT to MI of 10:1 to 1.67:1. However, there is a need for additional combinations of microbicides having enhanced activity against various strains of microorganisms to provide effective control of the 0 microorganisms. Moreover, there is a need for combinations containing lower levels of individual microbicides for environmental and economic benefit. The problem addressed by this invention is to provide such additional combinations of microbicides. A reference herein to a patent document or other matter which is given as 25 prior art is not to be taken as an admission that that document or matter was known or that the information it contains was part of the common general knowledge as at the priority date of any of the claims. Throughout the description and claims of the specification, the word "comprise" and variations of the word, such as "comprising" and "comprises", is 30 not intended to exclude other additives, components, integers or steps. 2123117.1 -2 STATEMENT OF THE INVENTION The present invention is directed to a microbicidal composition comprising: (a) N- (n-butyl)-1, 2-benzisothiazolin-3-one; and (b) 1,3-dimethylol-5,5 dimethyl hydantoin. 5 DETAILED DESCRIPTION OF THE INVENTION "BBIT" is N- (n-butyl) -1 ,2-benzisothiazolin- 3-one. "EDTA" is ethylenediamine tetraacetic acid. "MBIT" is N-methyl-1,2-benzisothiazolin-3 one. 0 As used herein, the following terms have the designated definitions, unless the context clearly indicates otherwise. The term "microbicide" refers to a compound capable of killing, inhibiting the growth of or controlling the growth of microorganisms at a locus; microbicides include bactericides, fungicides and algaecides. The term "microorganism" includes, for example, fungi (such as 5 yeast and mold), bacteria and algae. The term "locus" refers to an industrial system or product subject to contamination by microorganisms. The following abbreviations are used throughout the specification: ppm = parts per million by weight (weight/weight), mL = milliliter, ATCC = American Type Culture Collection, MBC = minimum biocidal concentration, and MIC = minimum 0 inhibitory concentration. Unless otherwise specified, temperatures are in degrees centigrade (*C), and references to percentages (%) are by weight. Amounts of organic microbicides are given on an active ingredient basis in ppm (w/w). The compositions of the present invention unexpectedly have been found 25 to provide enhanced microbicidal efficacy at a combined active ingredient level lower than that of the individual microbicides. In one embodiment of the invention, those antimicrobial compositions which contain halogenated 3 isothiazolones contain relatively low levels thereof, preferably no more than 1000 ppm, more preferably no more than 500 ppm, more preferably no more than 100 30 ppm, and most preferably no more than 50 ppm. Concentrations of halogenated 3-isothiazolones in the composition of this invention are based on the total weight of active ingredients in the composition, i.e., the microbicides exclusive of .3 any amounts of solvents, carriers, dispersants, stabilizers or other materials which may be present. In one embodiment of the invention, the antimicrobial composition contains less than 1000 ppm of 5-chloro-2-methyl-4-isothiazolin-3 one, more preferably no more than 500 ppm, more preferably no more than 100 5 ppm, and most preferably no more than 50 ppm. In one embodiment of the invention, the antimicrobial composition comprises N- (n-butyl) -1 ,2-benzisothiazolin- 3-one and 1,3-dimethylol-5,5 dimethylhydantoin. Preferably, a weight ratio of N-(n-butyl)-1,2 benzisothiazolin-3-one to 1,3-dimethylol-5,5-dimethylhydantoin is from 1:0.02 to 0 1:200, more preferably from 1:0.06 to 1:160. The microbicides in the composition of this invention may be used "as is" or may first be formulated with a solvent or a solid carrier. Suitable solvents include, for example, water; glycols, such as ethylene glycol, propylene glycol, diethylene glycol, dipropylene glycol, polyethylene glycol, and polypropylene 5 glycol; glycol ethers; alcohols, such as methanol, ethanol, propanol, phenethyl alcohol and phenoxypropanol; ketones, such as acetone and methyl ethyl ketone; esters, such as ethyl acetate, butyl acetate, triacetyl citrate, and glycerol triacetate; carbonates, such as propylene carbonate and dimethyl carbonate; and mixtures thereof. It is preferred that the solvent is selected from water, glycols, 0 glycol ethers, esters and mixtures thereof. Suitable solid carriers include, for example, cyclodextrin, silicas, diatomaceous earth, waxes, cellulosic materials, alkali and alkaline earth (e.g., sodium, magnesium, potassium) metal salts (e.g., chloride, nitrate, bromide, sulfate) and charcoal. When a microbicide component is formulated in a solvent, the formulation 25 may optionally contain surfactants. When such formulations contain surfactants, they are generally in the form of emulsive concentrates, emulsions, microemulsive concentrates, or microemulsions. Emulsive concentrates form emulsions upon the addition of a sufficient amount of water. Microemulsive concentrates form microemulsions upon the addition of a sufficient amount of 30 water. Such emulsive and microemulsive concentrates are generally well known in the art; it is preferred that such formulations are free of surfactants. U.S.

.4 Patent No. 5,444,078 may be consulted for further general and specific details on the preparation of various microemulsions and microemulsive concentrates. A microbicide component also can be formulated in the form of a dispersion. The solvent component of the dispersion can be an organic solvent or 5 water, preferably water. Such dispersions can contain adjuvants, for example, co-solvents, thickeners, anti-freeze agents, dispersants, fillers, pigments, surfactants, biodispersants, sulfosuccinates, terpenes, furanones, polycations, stabilizers, scale inhibitors and anti-corrosion additives. When both microbicides are each first formulated with a solvent, the D solvent used for the first microbicide may be the same as or different from the solvent used to formulate the other commercial microbicide, although water is preferred for most industrial biocide applications. It is preferred that the two solvents are miscible. Those skilled in the art will recognize that the microbicide components of 5 the present invention may be added to a locus sequentially, simultaneously, or may be combined before being added to the locus. It is preferred that the first microbicide and the second microbicide component be added to a locus simultaneously or sequentially. When the microbicides are added simultaneously or sequentially, each may individual components may contain 0 adjuvants, such as, for example, solvent, thickeners, anti-freeze agents, colorants, sequestrants (such as ethylenediamine-tetraacetic acid, ethylene diaminedisuccinic acid, iminodisuccinic acid and salts thereof), dispersants, surfactants, biodispersants, sulfosuccinates, terpenes, furanones, polycations, stabilizers, scale inhibitors and anti-corrosion additives. 2 5 The microbicidal compositions of the present invention can be used to inhibit the growth of microorganisms or higher forms of aquatic life (such as protozoans, invertebrates, bryozoans, dinoflagellates, crustaceans, mollusks, etc) by introducing a microbicidally effective amount of the compositions onto, into, or at a locus subject to microbial attack. Suitable loci include, for example: 30 industrial process water; electrocoat deposition systems,; cooling towers; air washers; gas scrubbers; mineral slurries; wastewater treatment; ornamental fountains; reverse osmosis filtration; ultrafiltration; ballast water; evaporative -5 condensers; heat exchangers; pulp and paper processing fluids and additives; starch; plastics; emulsions; dispersions; paints; latices; coatings, such as varnishes; construction products, such as mastics, caulks, and sealants; construction adhesives, such as ceramic adhesives, carpet backing adhesives, and 5 laminating adhesives; industrial or consumer adhesives; photographic chemicals; printing fluids; household products, such as bathroom and kitchen cleaners; cosmetics; toiletries; shampoos; soaps; detergents; industrial cleaners; floor polishes; laundry rinse water; metalworking fluids; conveyor lubricants; hydraulic fluids; leather and leather products; textiles; textile products; wood D and wood products, such as plywood, chipboard, wallboard, flakeboard, laminated beams, oriented strandboard, hardboard, and particleboard; petroleum processing fluids; fuel; oilfield fluids, such as injection water, fracture fluids, and drilling muds; agriculture adjuvant preservation; surfactant preservation; medical devices; diagnostic reagent preservation; food 5 preservation, such as plastic or paper food wrap; food, beverage, and industrial process pasteurizers; toilet bowls; recreational water; pools; and spas. Preferably, the microbicidal compositions of the present invention are used to inhibit the growth of microorganisms at a locus selected from one or more of mineral slurries, pulp and paper processing fluids and additives, starch, 0 emulsions, dispersions, paints, latices, coatings, construction adhesives, such as ceramic adhesives, carpet backing adhesives, photographic chemicals, printing fluids, household products such as bathroom and kitchen cleaners, cosmetics, toiletries, shampoos, soaps, detergents, industrial cleaners, floor polishes, laundry rinse water, metal working fluids, textile products, agriculture adjuvant ?5 preservation, surfactant preservation, diagnostic reagent preservation, food preservation, and food, beverage, and industrial process pasteurizers. The specific amount of the composition of this invention necessary to inhibit or control the growth of microorganisms and higher aquatic life forms in a locus depends upon the particular locus to be protected. Typically, the amount of 30 the composition of the present invention to control the growth of microorganisms in a locus is sufficient if it provides from 0.1 to 1,000 ppm of the isothiazoline ingredient of the composition in the locus. It is preferred that the isothiazolone -6 ingredients of the composition be present in the locus in an amount of at least 0.5 ppm, more preferably at least 4 ppm and most preferably at least 10 ppm. It is preferred that the isothiazolone ingredients of the composition be present in the locus in an amount of no more than 1000 ppm, more preferably no more than 500 5 ppm, and most preferably no more than 200 ppm. In one embodiment of the invention, the composition is substantially free of enzymatic biocides. Preferably, when BBIT or MBIT and either methylparaben or ethylparaben are combined, the composition is substantially free of enzymatic biocides. Enzymatic biocides are enzymes having activity 0 against microbes, as defined, e.g., in U.S. Pat. App. Pub. No. 2002/0028754. EXAMPLES Materials and Methods The synergism of the combination of the present invention was 5 demonstrated by testing a wide range of concentrations and ratios of the compounds. One measure of synergism is the industrially accepted method described by Kull, F.C.; Eisman, P.C.; Sylwestrowicz, H.D. and Mayer, R.L., in Applied Microbiology 9:538-541 (1961), using the ratio determined by the formula: 0 Qa/QA + Qb/QB = Synergy Index ("SI") wherein: QA = concentration of compound A (first component) in ppm, acting alone, which produced an end point (MIC of Compound A). 25 Qa = concentration of compound A in ppm, in the mixture, which produced an end point. QB = concentration of compound B (second component) in ppm, acting alone, which produced an end point (MIC of Compound B). Qb = concentration of compound B in ppm, in the mixture, which produced 30 an end point.

-7 When the sum of Qa/QA and Qb/QB is greater than one, antagonism is indicated. When the sum is equal to one, additivity is indicated, and when less than one, synergism is demonstrated. The lower the SI, the greater the synergy shown by that particular mixture. The minimum inhibitory concentration (MIC) 5 of a microbicide is the lowest concentration tested under a specific set of conditions that prevents the growth of added microorganisms. Synergy tests were conducted using standard microtiter plate assays with media designed for optimal growth of the test microorganism. Minimal salt medium supplemented with 0.2% glucose and 0.1% yeast extract (M9GY 0 medium) was used for testing bacteria; Potato Dextrose Broth (PDB medium) was used for testing yeast and mold. In this method, a wide range of combinations of microbicides and other personal care raw materials was tested by conducting high resolution MIC assays in the presence of various concentrations of BBIT or MBIT. High resolution MICs were determined by 5 adding varying amounts of microbicide to one column of a microtitre plate and doing subsequent ten-fold dilutions using an automated liquid handling system to obtain a series of endpoints ranging from 2 ppm to 10,000 ppm active ingredient. The synergy of the combinations of the present invention was determined 0 against a bacterium, Escherichia coli(E. coli- ATCC #8739), a yeast, Candida albicans (C albicans -- ATCC 10231), and a mold, Aspergillus niger (A. niger - ATCC 16404). The bacteria were used at a concentration of about 5 x 106 bacteria per mL and the yeast and mold at 5 x 101 fungi per mL. These microorganisms are representative of natural contaminants in many consumer 25 and industrial applications. The plates were visually evaluated for microbial growth (turbidity) to determine the MIC after various incubation times at 250 C (yeast and mold) or 300 C (bacteria). The test results for demonstration of synergy of the BBIT combinations of the present invention are shown below in Tables 1 through 26. In each test, First Component (A) was BBIT and the Second Component (B) was the other commercial microbicide. Each table shows the specific combinations of BBIT and the second component; results against the microorganisms tested with -8 incubation times; the end-point activity in ppm measured by the MIC for BBIT alone (QA), for the second component alone (Qs), for BBIT in the mixture (Qa) and for second component in the mixture (Qb); the calculated SI value; and the range of synergistic ratios for each combination tested (BBIT/second component or A/B). The test results for demonstration of synergy of the MBIT combinations of the present invention are shown below in Tables 27 through 54. In each test, First Component (A) was MBIT and the Second Component (B) was the other commercial microbicide. Each table shows the specific combinations of MBIT and the second component; results against the microorganisms tested with incubation times; the end-point activity in ppm measured by the MIC for MBIT alone (QA), for the second component alone (QB), for MBIT in the mixture (Qa) and for second component in the mixture (Qb); the calculated SI value; and the range of synergistic ratios for each combination tested (MBIT/second component or A/B). Table 1 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) 5 Second Component (B) = Benzalkonium chloride (Hyamine 3500) Microorganism Qa Qb SI A/B E coli8739 - M9GY 0 3 1.00 --.

(48 hours) 16.667 40 1.00 1/2 10 50 1.00 1/5 12.5 60 1.00 1/5 100 0 1.00 -- Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 60 1.00 -- (24 hours) 2 4 0.40 1/2 2 5 0.42 1/3 2 6 0.43 1/3 2 8 0.47 1/4 2 10 0.50 1/5 2 20 0.67 1/10 2 30 0.83 1/15 4 1 0.68 1/0.3 .9 Microorganism Qa Qb SI A/B 4 2 0.70 1/0.5 4 3 0.72 1/0.8 4 4 0.73 1/1 4 5 0.75 1/1 4 6 0.77 1/2 4 8 0.80 1/2 4 10 0.83 1/3 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 200 1.00 ---- (7 days) 5 60 0.55 1/12 5 80 0.65 1/16 5 100 0.75 1/20 10 50 0.75 1/5 10 60 0.80 1/6 10 80 0.90 1/8 15 4 0.77 1/0.3 15 5 0.78 1/0.3 15 6 0.78 1/0.4 15 8 0.79 1/0.5 15 10 0.80 1/0.7 15 20 0.85 1/1 15 30 0.90 1/2 20 0 1.00

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The ratios of BBIT/benzalkonium chloride tested ranged from 1/0.02 to 1/500. 5 Table 2 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Benzethonium chloride (Hyamine 1622) Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20 1.00 ---- (72 hours) 40 6 0.70 1/0.2 40 8 0.80 1/0.2 40 10 0.90 1/0.3 50 3 0.65 1/0.06 50 4 0.70 1/0.08 -10 Microorganism Qa Qb SI A/B 50 5 0.75 1/0.1 50 6 0.80 1/0.1 50 8 0.90 1/0.2 60 1 0.65 1/0.02 60 2 0.70 1/0.03 60 3 0.75 1/0.05 60 4 0.80 1/0.07 60 5 0.85 1/0.08 60 6 0.90 1/0.1 70 4 0.90 1/0.06 100 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 4 1.00 ---- (24 hours) 4 1 0.92 1/0.3 6 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 10 1.00 ---- (7 days) 5 4 0.65 1/0.8 5 5 0.75 1/0.8 5 6 0.85 1/1 10 4 0.90 1/0.4 20 0 1.00

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The ratios of BBIT/benzethonium chloride tested ranged from 1/0.02 to 1/500. 5 Table 3 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Benzyl alcohol 10 Microorganism Qa Qb SI A/B E. coli 8739 - M9GY 0 2000 1.00 ---- (24 hours) 100 2000 0.90 1/20 200 0 1.00 --- Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 3000 1.00 ---- (48 hours) 2.5 600 0.45 1/240 2.5 800 0.52 1/320 -11 Microorganism Qa Qb SI A/B 2.5 1000 0.58 1/400 2.5 2000 0.92 1/800 5 400 0.63 1/80 5 500 0.67 1/100 5 600 0.70 1/120 5 800 0.77 1/160 5 1000 0.83 1/200 7.5 20 0.76 1/3 7.5 30 0.76 1/4 7.5 40 0.76 1/5 7.5 50 0.77 1/7 7.5 60 0.77 1/8 7.5 80 0.78 1/11 7.5 100 0.78 1/13 7.5 200 0.82 1/27 7.5 300 0.85 1/40 7.5 400 0.88 1/53 7.5 500 0.92 1/67 7.5 600 0.95 1/80 10 0 1.00 --- Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 5000 1.00 ---- (3 days) 5 5000 1.13 1/1000 10 5000 1.25 1/500 15 4000 1.18 1/267 20 5000 1.50 1/250 40 0 1.00 --- The ratios of BBIT/benzyl alcohol tested ranged from 1/0.2 to 1/5000. 5 Table 4 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Benzisothiazolinone (BIT) Microorganism Qa Qb SI A/B E. coli 8739 - M9GY 0 8 1.00 ---- (72 hours) 40 1 0.53 1/0.03 40 2 0.65 1/0.05 -12 Microorganism Qa Qb SI A/B 40 3 0.78 1/0.08 40 4 0.90 1/0.1 50 1 0.63 1/0.02 50 2 0.75 1/0.4 50 3 0.88 1/0.06 60 1 0.73 1/0.02 60 2 0.85 1/0.03 100 0 1.00 --- Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 30 1.00 ---- (48 hours) 2.5 8 0.60 1/3 2.5 10 0.67 1/4 5 3 0.77 1/0.6 5 4 0.80 1/0.8 5 5 0.83 1/1 5 6 0.87 1/1.2 5 8 0.93 1/1.6 7.5 0 1.00 --- Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 100 1.00 ---- (7 days) 5 80 0.86 1/16 10 20 0.33 1/2 10 30 0.43 1/3 10 40 0.53 1/4 10 50 0.63 1/5 10 60 0.73 1/6 10 80 0.93 1/8 15 8 0.27 1/0.5 15 10 0.29 1/0.7 15 20 0.39 1/1 15 30 0.49 1/2 15 40 0.59 1/3 15 50 0.69 1/3 15 60 0.79 1/4 20 10 0.35 1/0.5 20 20 0.45 1/1 20 30 0.55 1/2 20 40 0.65 1/2 -13 Microorganism Qa Qb SI A/B 20 50 0.75 1/3 20 60 0.85 1/3 40 2 0.52 1/0.05 40 3 0.53 1/0.08 40 4 0.54 1/0.1 40 5 0.55 1/0.1 40 6 0.56 1/0.1 40 8 0.58 1/0.2 40 10 0.60 1/0.3 40 20 0.70 1/0.5 40 30 0.80 1/0.8 40 40 0.90 1/1 80 0 1.00 --- The ratios of BBIT/benzisothiazolinone tested ranged from 1/0.02 to 1/500. 5 Table.5 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = 2-bromo-2-nitro-propane-1,3-diol (BNPD) Microorganism Qa Qb SI A/B E. col 8739 - M9GY 0 10 1.00 ---- (48 hours) 40 1 0.50 1/0.03 40 2 0.60 1/0.05 40 3 0.70 1/0.08 40 4 0.80 1/0.1 40 5 0.90 1/0.1 50 1 0.60 1/0.02 50 2 0.70 1/0.04 50 3 0.80 1/0.06 50 4 0.90 1/0.08 70 1 0.80 1/0.01 70 2 0.90 1/0.03 80 1 0.90 1/0.01 100 0 1.00 --- Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 2000 1.00 ---- (48 hours) 2 1000 0.75 1/500 4 20 0.51 1/5 -14 Microorganism Qa Qb SI A/B 4 30 0.52 1/8 4 40 0.52 1/10 4 50 0.53 1/13 4 60 0.53 1/15 4 80 0.54 1/20 4 100 0.55 1/25 4 200 0.60 1/50 4 300 0.65 1/75 4 400 0.70 1/100 4 500 0.75 1/125 4 600 0.80 1/150 4 800 0.90 1/200 6 1 0.75 1/0.2 6 2 0.75 1/0.3 6 3 0.75 1/0.5 6 4 0.75 1/0.7 6 5 0.75 1/0.8 6 6 0.75 1/1 6 8 0.75 1/1 6 10 0.76 1/2 6 20 0.76 1/3 6 30 0.77 1/5 6 40 0.77 1/7 6 50 0.78 1/8 6 60 0.78 1/10 6 80 0.79 1/13 6 100 0.80 1/17 6 200 0.85 1/33 6 300 0.90 1/50 8 0 1.00 --- Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 2000 1.00 ---- (7 days) 5 1000 0.75 1/200 10 8 0.50 1/0.8 10 10 0.51 1/1 10 20 0.51 1/2 10 30 0.52 1/3 10 40 0.52 1/4 10 50 0.53 1/5 - 15 Microorganism Qa Qb SI A/B 10 60 0.53 1/6 10 80 0.54 1/8 10 100 0.55 1/10 10 200 0.60 1/20 10 300 0.65 1/30 10 400 0.70 1/40 10 500 0.75 1/50 10 600 0.80 1/60 10 800 0.90 1/80 20 0 1.00 --- The ratios of BBIT/2-bromo-2-nitro-propane-1,3-diol tested ranged from 1/0.01 to 1/500. 5 Table 6 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Butylene glycol 0 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20000 1.00 ---- (48 hours) 40 5000 0.65 1/125 40 6000 0.70 1/150 40 8000 0.80 1/200 40 10000 0.90 1/250 50 2000 0.60 1/40 50 3000 0.65 1/60 50 4000 0.70 1/80 50 5000 0.75 1/100 50 6000 0.80 1/120 50 8000 0.90 1/160 70 2000 0.80 1/29 70 3000 0.85 1/43 70 4000 0.90 1/57 80 200 0.81 1/3 80 300 0.82 1/4 80 400 0.82 1/5 80 500 0.83 1/6 80 600 0.83 1/8 80 800 0.84 1/10 -16 Microorganism Qa Qb SI A/B 80 1000 0.85 1/13 80 2000 0.90 1/25 100 0 1.00 --- Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 20000 1.00 ---- (24 hours) 2 10000 0.83 1/5000 4 200 0.68 1/50 4 300 0.68 1/75 4 400 0.69 1/100 4 500 0.69 1/125 4 600 0.70 1/150 4 800 0.71 1/200 4 1000 0.72 1/250 4 2000 0.77 1/500 4 3000 0.82 1/750 4 4000 0.87 1/1000 4 5000 0.92 1/1250 6 0 1.00 --- Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 10000 1.00 --- (3 days) 5 5000 0.83 5 6000 0.93 15 0 1.00 --- The ratios of BBIT/butylene glycol tested ranged from 1/0.2 to 1/5000. 5 Table 7 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B)= Caprylyl glycol Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 2000 1.00 ---- (24 hours) 40 400 0.60 1/10 40 500 0.65 1/13 40 600 0.70 1/15 40 800 0.80 1/20 40 1000 0.90 1/25 50 200 0.60 1/4 - 17 Microorganism Qa Qb SI A/B 50 600 0.80 1/12 50 800 0.90 1/16 70 200 0.80 1/3 70 300 0.85 1/4 70 400 0.90 1/6 80 200 0.90 1/3 100 0 1.00 --- Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 3000 1.00 ---- (24 hours) 2 2000 0.92 1/1000 4 500 0.67 1/125 4 600 0.70 1/150 4 800 0.77 1/200 4 1000 0.83 1/250 6 40 0.76 1/7 6 50 0.77 1/8 6 60 0.77 1/10 6 80 0.78 1/13 6 100 0.78 1/17 6 200 0.82 1/33 6 300 0.85 1/50 6 400 0.88 1/67 6 500 0.92 1/83 8 0 1.00

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A. niger 16404 - PDB 0 3000 1.00 ---- (7 days) 5 1000 0.58 1/200 5 2000 0.92 1/400 10 1000 0.83 1/100 20 0 1.00 ---- The ratios of BBIT/caprylyl glycol tested ranged from 1/0.2 to 1/5000. 5 Table 8 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Chlorphenesin Microorganism Qa Qb SI A/B E. col 8739 - M9GY 0 2000 1.00 ----- - 18 Microorganism Qa Qb SI A/B (24 hours) 40 500 0.65 1/13 40 600 0.70 1/115 40 800 0.80 1/20 40 1000 0.90 1/25 50 300 0.65 1/6 50 400 0.70 1/8 50 500 0.75 1/10 50 600 0.80 1/12 50 800 0.90 1/16 70 200 0.80 1/3 70 300 0.85 1/4 70 400 0.90 1/6 80 100 0.85 1/1 80 200 0.90 1/3 100 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 2000 1.00 ---- (24 hours) 2 800 0.65 1/400 2 1000 0.75 1/500 4 300 0.65 1/75 4 400 0.70 1/100 4 500 0.75 1/125 4 600 0.80 1/150 4 800 0.90 1/200 6 10 0.76 1/2 6 20 0.76 1/3 6 30 0.77 1/5 6 40 0.77 1/7 6 50 0.78 1/8 6 60 0.78 1/10 6 80 0.79 1/13 6 100 0.80 1/17 6 200 0.85 1/33 6 300 0.90 1/50 8 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 2000 1.00 ---- (4 days) 5 1000 0.83 1/200 -19 Microorganism Qa Qb SI A/B 15 0 1.00

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The ratios of BBIT/chlorphenesin tested ranged from 1/0.2 to 1/5000. 5 Table 9 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B)= DMDM Hydantoin (DMDMH) Microorganism Q. Qi SI A/B E. colj8739 - M9GY 0 200 1.00 ---- (72 hours) 40 8 0.44 1/0.2 40 10 0.45 1/0.3 40 20 0.50 1/0.5 40 30 0.55 1/0.8 40 40 0.60 1/1 40 50 0.65 1/1 40 60 0.70 1/2 40 80 0.80 1/2 40 100 0.90 1/3 50 3 0.52 1/0.06 50 4 0.52 1/0.08 50 5 0.53 1/0.1 50 6 0.53 1/0.1 50 8 0.54 1/0.2 50 10 0.55 1/0.2 50 20 0.60 1/0.4 50 30 0.65 1/0.6 50 40 0.70 1/0.8 50 50 0.75 1/1 50 60 0.80 1/1.2 50 80 0.90 1/1.6 60 6 0.63 1/0.1 60 8 0.64 1/0.1 60 10 0.65 1/0.2 60 20 0.70 1/0.3 60 30 0.75 1/0.5 60 40 0.80 1/0.7 60 50 0.85 1/0.8 -20 Microorganism Qa Qb SI A/B 60 60 0.90 1/1 80 8 0.84 1/0.1 80 10 0.85 1/0.1 80 20 0.90 1/0.3 100 0 1.00

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C albicans 10231 - PDB 0 3000 1.00 ---- (24 hours) 5 600 0.87 1/120 5 800 0.93 1/160 7.5 0 1.00

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Microorganism Q. Qb SI A/B A. niger 16404 - PDB 0 1000 1.00 ---- (7 days) 20 80 0.33 1/4 20 100 0.35 1/5 20 200 0.45 1/10 20 300 0.55 1/15 20 400 0.65 1/20 20 500 0.75 1/25 20 600 0.85 1/30 40 100 0.60 1/3 40 200 0.70 1/5 40 300 0.80 1/8 40 400 0.90 1/10 80 0 1.00

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The ratios of BBIT/DMDM Hydantoin tested ranged from 1/0.2 to 1/5000. 5 Table 10 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = dithio-2, 2'-bis(N-methylbenzamide) (DTBMA) Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 60 1.00 ---- (48 hours) 40 6 0.50 1/0.2 40 8 0.53 1/0.2 40 10 0.57 1/0.3 40 20 0.73 1/0.5 40 30 0.90 1/0.8 50 3 0.55 1/1.06 50 4 0.57 1/0.08 -21 Microorganism Q. Qb SI A/B 50 5 0.58 1/0.1 50 6 0.60 1/0.1 50 8 0.63 1/1.2 50 10 0.67 1/0.2 50 20 0.83 1/0.4 60 3 0.65 1/0.05 60 4 0.67 1/0.07 60 5 0.68 1/008 60 6 0.70 1/0.1 60 8 0.73 1/0.1 60 10 0.77 1/0.2 60 20 0.93 1/0.3 70 3 0.75 1/0.04 70 4 0.77 1/0.06 70 5 0.78 1/0.07 70 6 0.80 1/0.09 70 7 0.82 1/0.1 70 8 0.83 1/0.1 70 10 0.87 1/0.1 80 1 0.82 1/0.01 80 2 0.83 1/0.03 80 3 0.85 1/0.04 80 4 0.87 1/0.05 80 5 0.88 1/0.06 80 6 0.90 1/0.08 80 8 0.93 1/0.1 100 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 2000 1.00 ---- (24 hours) 8 40 0.55 1/5 8 50 0.56 1/6 8 60 0.56 1/8 8 80 0.57 1/10 8 100 0.58 1/13 8 200 0.63 1/13 8 300 0.68 1/38 8 400 0.73 1/50 8 500 0.78 1/63 -22 Microorganism Qa Qb SI A/B 8 600 0.83 1/75 8 800 0.93 1/100 10 1 0.67 1/0.1 10 2 0.67 1/0.2 10 3 0.67 1/0.3 10 4 0.67 1/0.4 10 5 0.67 1/0.5 10 6 0.67 1/0.6 10 8 0.67 1/0.8 10 10 0.67 1/1 10 20 0.68 1/2 10 30 0.68 1/3 10 40 0.69 1/4 10 50 0.69 1/5 10 60 0.70 1/6 10 80 0.71 1/8 10 100 0.72 1/10 10 200 0.77 1/20 10 300 0.82 1/30 10 400 0.87 1/40 10 500 0.92 1/50 15 0 1.00

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A. niger 16404 - PDB 0 1000 1.00 ---- (7 days) 5 800 0.93 1/160 40 0 1.00

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The ratios of BBIT/ dithio-2, 2'-bis(N-methylbenzamide) tested ranged from 1/0.02 to 1/500. 5 Table 11 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Ethylenediamine tetraacetic acid (EDTA) 10 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 40000 1.00 ---- (72 hours) 40 800 0.42 1/20 40 1000 0.43 1/25 40 1200 0.43 1/30 40 1600 0.44 1/40 40 2000 0.45 1/50 -23 Microorganism Qa Qb SI A/B 40 4000 0.50 1/100 40 6000 0.55 1/150 40 8000 0.60 1/200 40 10000 0.65 1/250 40 12000 0.70 1/300 40 16000 0.80 1/400 40 20000 0.90 1/500 50 400 0.51 1/8 50 600 0.52 1/12 50 800 0.52 1/16 50 1000 0.53 1/20 50 1200 0.53 1/24 50 1600 0.54 1/32 50 2000 0.55 1/40 50 4000 0.60 1/80 50 6000 0.65 1/120 50 8000 0.70 1/160 50 10000 0.75 1/200 50 12000 0.80 1/240 50 16000 0.90 1/320 60 400 0.61 1/7 60 600 0.62 1/10 60 800 0.62 1/13 60 1000 0.63 1/17 60 1200 0.63 1/20 60 1600 0.64 1/27 60 2000 0.65 1/33 60 4000 0.70 1/67 60 6000 0.75 1/100 60 8000 0.80 1/133 60 10000 0.85 1/167 60 12000 0.90 1/200 70 200 0.71 1/3 70 400 0.71 1/6 70 600 0.72 1/9 70 800 0.72 1/11 70 1000 0.73 1/14 70 1200 0.73 1/17 -24 Microorganism Qa Qb SI A/B 70 1600 0.74 1/23 70 2000 0.75 1/29 70 4000 0.80 1/57 70 6000 0.85 1/86 70 8000 0.90 1/114 70 10000 0.95 1/143 80 400 0.81 1/5 80 600 0.82 1/8 80 800 0.82 1/40 80 1000 0.83 1/13 80 1200 0.83 1/15 80 1600 0.84 1/20 80 2000 0.85 1/25 80 4000 0.90 1/50 80 6000 0.95 1/75 100 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 600 1.00 ---- (24 hours) 2.5 60 0.43 1/24 2.5 80 0.47 1/32 2.5 100 0.50 1/40 2.5 120 0.53 1/48 2.5 160 0.60 1/64 2.5 200 0.67 1/80 5 60 0.77 1/12 5 80 0.80 1/16 5 100 0.83 1/20 5 120 0.87 1/24 5 160 0.93 1/32 7.5 0 1.00

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A. niger 16404 - PDB 0 1600 1.00 ---- (7 days) 5 1000 0.75 1/200 5 1200 0.88 1/240 10 400 0.50 1/40 10 600 0.63 1/60 10 800 0.75 1/80 10 1000 0.88 1/100 15 400 0.63 1/27 15 600 0.75 1/40 15 800 0.88 1/53 20 200 0.63 1/10 -25 Microorganism Qa Qb SI A/B 20 400 0.75 1/20 20 600 0.88 1/30 40 0 1.00

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The ratios of BBIT/ethylenediamine tetraacetic acid tested ranged from 1/0.4 to 1/10000. 5 Table 12 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B)= Ethylparaben 0 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 2000 1.00 ---- (24 hours) 60 1000 0.80 1/17 80 800 0.80 1/10 80 1000 0.90 1/13 100 200 0.60 1/2 100 300 0.65 1/3 100 400 0.70 1/4 100 500 0.75 1/5 100 600 0.80 1/6 100 800 0.90 1/8 200 0 1.00

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Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 800 1.00 ---- (24 hours) 2.5 80 0.43 1/32 2.5 100 0.46 1/40 2.5 200 0.58 1/80 2.5 300 0.71 1/120 2.5 400 0.83 1/160 2.5 500 0.96 1/200 5 30 0.70 1/6 5 40 0.72 1/8 5 50 0.73 1/10 5 60 0.74 1/12 5 80 0.77 1/16 5 100 0.79 1/20 5 200 0.92 1/40 -26 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 800 1.00 -.

(7 days) 15 600 0.94 1/40 80 0 1.00

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The ratios of BBIT/ethylparaben tested ranged from 1/0.2 to 1/5000. Table 13 5 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Hexamidine diisethionate Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 100 1.00 -- (48 hours) 40 1 0.57 1/0.03 40 2 0.73 1/0.05 40 3 0.90 1/0.08 50 1 0.67 1/0.02 50 2 0.83 1/0.04 60 1 0.77 1/0.02 60 2 0.93 1/0.03 70 1 0.87 1/0.01 80 1 0.97 1/0.01 6 0 1.00 -- Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 10 1.00 - (24 hours) 2 4 0.60 1/2 2 5 0.70 1/2.5 2 6 0.80 1/3 4 2 0.60 1/0.5 4 3 0.70 1/0.75 4 4 0.80 1/1 4 5 0.90 1/1.3 6 1 0.70 1/0.2 6 2 0.80 1/0.3 6 3 0.90 1/0.5 8 1 0.90 1/1.1 10 0 1.00 --- -27 The ratios of BBIT/hexamidine diisethionate tested ranged from 1/0.02 to 1/500. 5 Table 14 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Hexetidine Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 200 1.00 ---- (72 hours) 40 10 0.33 1/0.3 40 20 0.45 1/1.5 40 30 0.58 1/0.8 40 40 0.70 1/1 40 50 0.83 1/1.3 40 60 0.95 1/1.5 50 10 0.38 1/0.2 50 20 0.50 1/0.4 50 30 0.63 1/0.6 50 40 0.75 1/0.8 50 50 0.88 1/1 60 4 0.35 1/0.07 60 5 0.36 1/0.08 60 6 0.38 1/0.1 60 8 0.40 1/0.1 60 10 0.43 1/0.2 60 20 0.55 1/0.3 60 30 0.68 1/0.5 60 40 0.80 1/0.7 60 50 0.93 1/0.8 70 5 0.41 1/0.07 70 6 0.43 1/0.09 70 8 0.45 1/0.1 70 10 0.48 1/0.1 70 20 0.60 1/0.3 70 30 0.73 1/0.4 70 40 0.85 1/0.6 70 50 0.98 1/0.7 80 4 0.45 1/0.05 80 5 0.46 1/0.06 -28 Microorganism Qa Qb SI A/B 80 6 0.48 1/0.08 80 8 0.50 1/0.1 80 10 0.53 1/0.1 80 20 0.65 1/0.3 80 30 0.78 1/0.4 80 40 0.90 1/0.5 100 1 0.51 1/0.01 100 2 0.53 1/0.02 100 3 0.54 1/0.03 100 4 0.55 1/0.04 100 5 0.56 1/0.05 100 6 0.58 1/0.06 100 8 0.60 1/0.08 100 10 0.63 1/0.1 100 20 0.75 1/0.2 100 30 0.88 1/0.3 80 0 1.00

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Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 100 1.00 ---- (72 hours) 2 40 0.53 1/20 2 50 0.63 1/25 2 60 0.73 1/30 2 80 0.93 1/40 4 30 0.57 1/8 4 40 0.67 1/10 4 50 0.77 1/13 4 60 0.87 1/15 6 20 0.60 1/3 6 30 0.70 1/5 6 40 0.80 1/7 6 50 0.90 1/9 8 5 0.58 1/0.6 8 6 0.59 1/0.8 8 8 0.61 1/1 8 10 0.63 1/1 8 20 0.73 1/3 8 30 0.83 1/4 8 40 0.93 1/5 10 1 0.68 1/0.1 10 2 0.69 1/0.2 -29 Microorganism Qa Qb SI AB 10 3 0.70 1/0.3 10 4 0.71 1/0.4 10 5 0.72 1/0.5 10 6 0.73 1/0.6 10 8 0.75 1/0.8 10 10 0.77 1/1 10 20 0.87 1/2 10 30 0.97 1/3 15 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 100 1.00 ---- (4 days) 5 50 0.75 1/10 5 60 0.85 1/12 10 40 0.90 1/4 20 0 1.00

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The ratios of BBIT/hexetidine tested ranged from 1/0.02 to 1/500. 5 Table 15 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Hexylene glycol 0 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20000 1.00 ---- (24 hours) 40 20000 1.40 1/500 50 20000 1.50 1/400 60 20000 1.60 1/333 80 20000 1.80 1/250 100 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 20000 1.00 ---- (24 hours) 2 20000 1.20 1/10000 4 20000 1.40 1/5000 6 20000 1.60 1/3333 8 20000 1.80 1/2800 10 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 20000 1.00 ----- - 30 Microorganism Qa Qb SI A/B (3 days) 5 20000 1.33 1/4000 10 20000 1.67 1/2000 15 0 1.00

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The ratios of BBIT/hexylene glycol tested ranged from 1/0.2 to 1/5000. No synergy was observed between BBIT and hexylene glycol. 5 Table 16 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Methylparaben 0 Microorganism Qa Qb SI A/B E coli 8739 - M9GY 0 2000 1.00 ---- (72 hours) 40 400 0.60 1/10 40 500 0.65 1/13 40 600 0.70 1/15 40 800 0.80 1/20 40 1000 0.90 1/25 50 100 0.55 1/2 50 200 0.60 1/4 50 300 0.65 1/6 50 400 0.70 1/8 50 500 0.75 1/10 50 600 0.80 1/12 50 800 0.90 1/16 60 30 0.62 1/0.5 60 40 0.62 1/0.7 60 50 0.63 1/0.8 60 60 0.63 1/1 60 80 0.64 1/1 60 100 0.65 1/2 60 200 0.70 1/3 60 300 0.75 1/5 60 400 0.80 1/7 60 500 0.85 1/8 60 600 0.90 1/10 70 300 0.85 1/4 70 400 0.90 1/6 -31 Microorganism Qa Qb SI A/B 70 500 0.95 1/7 80 300 0.95 1/4 100 0 1.00

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Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 1000 1.00 ---- (24 hours) 2.5 100 0.43 1/40 2.5 200 0.53 1/80 2.5 300 0.63 1/120 2.5 400 0.73 1/160 2.5 500 0.83 1/200 2.5 600 0.93 1/240 5 40 0.71 1/8 5 50 0.72 1/10 5 60 0.73 1/12 5 80 0.75 1/16 5 100 0.77 1/20 5 200 0.87 1/40 5 300 0.97 1/60 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 1000 1.00 ---- (3 days) 5 600 0.73 1/120 5 800 0.93 1/160 40 0 1.00

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The ratios of BBIT/methylparaben tested ranged from 1/0.2 to 1/5000. 5 Table 17 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Pentylene glycol Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20000 1.00 ---- (24 hours) 40 20000 1.40 1/500 60 20000 1.60 1/333 80 20000 1.80 1/250 100 0 1.00

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Microorganism Qa Qb SI A/B -32 Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 20000 1.00 ---- (24 hours) 2 20000 1.02 1/10000 4 20000 1.04 1/5000 8 20000 1.08 10 20000 1.10 15 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 20000 1.00 ---- (4 days) 5 20000 1.05 10 20000 1.10 15 20000 1.15 20 0 1.00

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The ratios of BBIT/pentylene glycol tested ranged from 1/0.2 to 1/5000. No synergy was observed between BBIT and pentylene glycol. 5 Table 18 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Phenoxyethanol 0 Microorganism Qa Qb SI A/B E. co/i8739 - M9GY 0 5000 1.00 ---- (72 hours) 50 800 0.66 1/16 50 1000 0.70 1/20 50 2000 0.90 1/40 60 60 0.61 1/1 60 80 0.62 1/1 60 100 0.62 1/2 60 200 0.64 1/3 60 300 0.66 1/5 60 400 0.68 1/7 60 500 0.70 1/8 60 600 0.72 1/10 60 800 0.76 1/13 60 1000 0.80 1/17 70 1000 0.90 1/14 100 0 1.00

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Microorganism Qa Qb SI A/B - 33 Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 4000 1.00 ---- (24 hours) 2.5 1000 0.58 1/400 2.5 2000 0.83 1/800 5 500 0.79 1/100 5 600 0.82 1/120 5 800 0.87 1/160 5 1000 0.92 1/200 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 4000 1.00 -- (7 days) 5 3000 0.88 1/600 40 0 1.00 -- The ratios of BBIT/phenoxyethanol tested ranged from 1/0.2 to 1/5000. 5 Table 19 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B)= linoleamidopropyl PG-dimonium chloride phosphate (Phospholipid EFA) 0 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 600 1.00 -- (48 hours) 40 400 0.87 1/10 50 400 0.92 1/8 200 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 2000 1.00 ---- (24 hours) 2 800 0.60 1/400 2 1000 0.70 1/500 4 600 0.70 1/150 4 800 0.80 1/200 4 1000 0.90 1/250 6 500 0.85 1/83 6 600 0.90 1/100 10 0 1.00 -- Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 2000 1.00 - (4 days) 5 600 0.55 1/120 -34 Microorganism Qa Qb SI A/B 5 800 0.65 1/160 5 1000 0.75 1/200 10 600 0.80 1/60 10 800 0.90 1/80 20 0 1.00

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The ratios of BBIT/linoleamidopropyl PG-dimonium chloride phosphate tested ranged from 1/0.02 to 1/500. 5 Table 20 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = cocamidopropyl PG-dimonium chloride phosphate 0 (Phospholipid PTC) Microorganism Qa Q SI A/B E. coli 8739 - M9GY 0 80 1.00 ---- (72 hours) 40 20 0.45 1/0.5 40 30 0.58 1/0.75 40 40 0.70 1/1 40 50 0.83 1/0.3 40 60 0.95 1/2 50 20 0.50 1/0.4 50 30 0.63 1/0.6 50 40 0.75 1/0.8 50 50 0.88 1/1 60 8 0.40 1/0.1 60 10 0.43 1/0.2 60 20 0.55 1/0.3 60 30 0.68 1/0.5 60 40 0.80 1/0.7 60 50 0.93 1/0.8 70 10 0.48 1/0.1 70 20 0.60 1/0.3 70 30 0.73 1/0.4 70 40 0.85 1/0.6 70 50 0.98 1/0.7 80 10 0.53 1/0.1 -35 Microorganism Q. Qb SI A/B 80 20 0.65 1/0.3 80 30 0.78 1/0.4 80 40 0.90 1/0.5 100 3 0.54 1/0.3 100 4 0.55 1/0.4 100 5 0.56 1/0.5 100 6 0.58 1/0.6 100 8 0.60 1/0.8 100 10 0.63 1/0.1 100 20 0.75 1/0.2 100 30 0.88 1/0.3 200 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 1000 1.00 ---- (24 hours) 2 600 0.73 1/300 2 800 0.93 1/400 4 400 0.67 1/400 4 500 0.77 1/125 4 600 0.87 1/150 6 300 0.70 1/50 6 400 0.80 1/67 6 500 0.90 1/83 8 4 0.54 1/0.5 8 5 0.54 1/0.6 8 6 0.54 1/0.8 8 8 0.54 1/1 8 10 0.54 1/1 8 20 0.55 1/6 8 30 0.56 1/4 8 40 0.57 1/5 8 50 0.58 1/6 8 60 0.59 1/8 8 80 0.61 1/10 8 100 0.63 1/13 8 200 0.73 1/25 8 300 0.83 1/38 -36 Microorganism Qa Qb SI A/B 8 400 0.93 1/50 10 3 0.67 1/0.3 10 4 0.67 1/0.4 10 5 0.67 1/0.5 10 6 0.67 1/0.6 10 8 0.67 1/0.8 10 10 0.68 1/1 10 20 0.69 1/2 10 30 0.70 1/3 10 40 0.71 1/4 10 50 0.72 1/5 10 60 0.73 1/6 10 80 0.75 1/8 10 100 0.77 1/10 10 200 0.87 1/20 10 300 0.97 1/30 15 0 1.00 ---- Microorganism Q. Qb SI A/B A. niger 16404 - PDB 0 1000 1.00 ---- (3 days) 5 500 0.83 1/100 5 600 0.93 1/120 10 200 0.87 1/20 10 300 0.97 1/30 15 0 1.00 ---- The ratios of BBIT/cocamidopropyl PG-dimonium chloride phosphate tested ranged from 1/0.02 to 1/500. 5 Table 21 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Potassium sorbate L0 Microorganism Qa Qb SI A/B E. coli 8739 - M9GY 0 20000 1.00 ---- (24 hours) 20 20000 1.10 1/1000 40 20000 1.20 1/500 60 20000 1.30 1/333 - 37 Microorganism Qa Qb SI A/B 80 20000 1.40 1/250 100 20000 1.50 1/200 200 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 400 1.00 ---- (24 hours) 2.5 100 0.58 1/40 2.5 200 0.83 1/80 5 30 0.74 1/6 5 60 0.82 1/12 5 80 0.87 1/16 5 100 0.92 1/20 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 8000 1.00 ---- (3 days) 5 6000 0.88 1/1200 10 4000 0.75 1/400 10 5000 0.88 1/500 15 4000 0.88 1/267 20 600 0.58 1/30 20 800 0.60 1/40 20 1000 0.63 1/50 20 2000 0.75 1/100 20 3000 0.88 1/150 40 0 1.00

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The ratios of BBIT/potassium sorbate tested ranged from 1/0.2 to 1/5000. 5 Table 22 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Propylparaben Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 2000 1.00 ---- (24 hours) 20 2000 1.10 1/100 40 2000 1.20 1/50 60 2000 1.30 1/33 80 2000 1.40 1/25 100 2000 1.50 1/25 - 38 Microorganism Qa Qb SI A/B 200 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 2000 1.00 ---- (24 hours) 2.5 30 0.35 1/12 2.5 40 0.35 1/16 2.5 50 0.36 1/20 2.5 60 0.36 1/24 2.5 80 0.37 1/32 2.5 100 0.38 1/40 2.5 200 0.43 1/80 2.5 300 0.48 1/120 2.5 400 0.53 1/160 2.5 500 0.58 1/200 2.5 600 0.63 1/240 2.5 800 0.73 1/320 2.5 1000 0.83 1/400 5 20 0.68 1/4 5 30 0.68 1/6 5 40 0.69 1/8 5 50 0.69 1/10 5 60 0.70 1/12 5 80 0.71 1/16 5 100 0.72 1/20 5 200 0.77 1/40 5 300 0.82 1/60 5 400 0.87 1/80 5 500 0.92 1/100 5 600 0.97 1/120 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 2000 1.00 ---- (7 days) 10 1000 0.63 1/100 20 800 0.65 1/40 20 1000 0.75 1/80 40 800 0.90 1/20 80 0 1.00

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The ratios of BBIT/propylparaben tested ranged from 1/0.2 to 1/5000.

- 39 Table 23 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) 5 Second Component (B) = Quaternium-15 (active - 1-(3-chloroallyl)-3,5,7-triaza-l azoniaadamantane chloride) Microorganism Qa Qb SI A/B E. col 8739 - M9GY 0 200 1.00 ---- (72 hours) 40 50 0.43 1/1 40 60 0.43 1/1 40 80 0.44 1/2 40 100 0.45 1/3 40 200 0.50 1/5 40 300 0.55 1/8 40 400 0.60 1/10 40 500 0.65 1/13 40 600 0.70 1/15 40 800 0.80 1/20 200 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 600 1.00 ---- (24 hours) 5 50 0.75 1/10 5 60 0.77 1/12 5 80 0.80 1/16 5 100 0.83 1/20 7.5 0 1.00

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Microorganism Q. Qb SI A/B A. niger 16404 - PDB 0 3000 1.00 ---- (4 days) 10 1000 0.58 1/100 10 2000 0.92 1/200 20 1000 0.83 1/50 40 0 1.00

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The ratios of BBIT/1-(3-chloroallyl)-3,5,7-triaza-1-azoniaadamantane 10 chloride tested ranged from 1/0.02 to 1/5000.

- 40 Table 24 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) 5 Second Component (B) = dehydroacetic acid, sodium salt (SDHA) Microorganism Qa Qb SI A/B E coli8739 - M9GY 0 20000 1.00 (24 hours) 20 20000 1.10 1/1000 40 20000 1.20 1/500 60 20000 1.30 1/333 80 20000 1.40 1/250 100 20000 1.50 1/200 200 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 40 1.00 (24 hours) 4 10 0.75 1/3 6 4 0.85 1/0.7 6 5 0.88 1/0.8 6 6 0.90 1/1 6 8 0.95 1/1 8 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 200 1.00 ---- (7 days) 20 80 0.65 1/4 20 100 0.75 1/5 40 50 0.75 1/3 40 60 0.80 1/2 40 80 0.90 1/2 80 0 1.00

----

The ratios of BBIT/dehydroacetic acid, sodium salt tested ranged from 1/0.02 to 1/5000. 10 Table 25 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) 15 Second Component (B) = Sodium benzoate Microorganism Qa Qb SI A/B E coli 8739 - M9GY 0 20000 1.00 ----- -41 Microorganism Qa Qb SI A/B (24 hours) 20 20000 1.10 1/1000 40 20000 1.20 1/500 60 20000 1.30 1/333 80 20000 1.40 1/250 100 20000 1.50 1/200 200 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 2000 1.00 ---- (48 hours) 2.5 400 0.53 1/160 2.5 500 0.58 1/200 2.5 600 0.63 1/240 2.5 800 0.73 1/320 2.5 1000 0.83 1/400 5 300 0.82 1/60 5 400 0.87 1/80 5 500 0.92 1/100 5 600 0.97 1/120 7.5 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 10000 1.00 ---- (7 days) 5 8000 0.86 1/1600 10 3000 0.43 1/300 10 4000 0.53 1/400 10 5000 0.63 1/500 10 6000 0.73 1/600 10 8000 0.93 1/800 15 3000 0.49 1/200 15 4000 0.59 1/267 15 5000 0.69 1/333 15 6000 0.79 1/400 15 7000 0.89 1/467 15 8000 0.99 1/533 20 1000 0.35 1/50 20 2000 0.45 1/100 20 3000 0.55 1/150 20 4000 0.65 1/200 20 5000 0.75 1/250 20 6000 0.85 1/300 -42 Microorganism Qa Qb SI A/B 20 7000 0.95 1/350 40 500 0.55 1/13 40 600 0.56 1/15 40 800 0.58 1/20 40 1000 0.60 1/25 40 2000 0.70 1/50 40 3000 0.80 1/75 40 4000 0.90 1/100 80 0 1.00

----

The ratios of BBIT/sodium benzoate tested ranged from 1/0.2 to 1/5000. 5 Table 26 First Component (A) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Second Component (B) = Sodium citrate Microorganism Q. Qb SI A/B E coli8739 - M9GY 0 20000 1.00 ---- (24 hours) 20 20000 1.10 1/1000 40 20000 1.20 1/500 60 20000 1.30 1/333 80 20000 1.40 1/250 100 20000 1.50 1/200 200 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 10000 1.00 ---- (48 hours) 2.5 3000 0.55 1/1200 2.5 4000 0.65 1/1600 2.5 5000 0.75 1/2000 2.5 6000 0.85 1/2400 5 600 0.56 1/120 5 800 0.58 1/160 5 1000 0.60 1/200 5 2000 0.70 1/400 5 3000 0.80 1/600 5 4000 0.90 1/800 10 0 1.00

----

Microorganism Qa Qb SI A/B -43 Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 10000 1.00 ---- (3 days) 5 8000 0.93 1/1600 10 5000 0.75 1/500 10 6000 0.85 1/600 15 3000 0.68 1/200 15 4000 0.78 1/267 15 5000 0.88 1/333 15 6000 0.98 1/400 20 1000 0.60 1/50 20 2000 0.70 1/100 20 3000 0.80 1/150 20 4000 0.90 1/200 40 0 1.00

----

The ratios of BBIT/sodium citrate tested ranged from 1/0.2 to 1/5000. 5 Table 27 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = N-(n-butyl)-1,2-benzisothiazolinone (BBIT) Microorganism Qa Qb SI A/B E coli8739 - M9GY 0 100 1.00 ---- (24 hours) 2.5 50 0.75 1/20 2.5 60 0.85 1/24 5 40 0.90 1/8 7.5 10 0.85 1/1 7.5 20 0.95 1/3 10 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 6 1.00 ---- (48 hours) 2.5 3 0.75 1/1 2.5 4 0.92 1/2 10 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 40 1.00 ---- (4 days) 10 5 0.38 1/0.5 10 6 0.40 1/0.6 -44 Microorganism Qa Qb SI A/B 10 8 0.45 1/0.8 10 10 0.50 1/1 10 20 0.75 1/2 20 6 0.65 1/0.3 20 8 0.70 1/0.4 20 10 0.75 1/0.5 40 0 1.00

----

The ratios of MBIT/BBIT tested ranged from 1/0.01 to 1/400. 5 Table 28 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Benzalkonium chloride (Hyamine 3500) Microorganism Qa Qb SI A/B E. coli 8739 - M9GY 0 10 1.00 ---- (24 hours) 2.5 4 1.25 1/2 5 3 1.25 1/0.6 7.5 3 1.50 1/0.4 100 0 1.00

----

Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 60 1.00 ---- (24 hours) 2 10 0.42 1/5 2 20 0.58 1/10 2 30 0.75 1/15 2 40 0.92 1/20 4 3 0.55 1/0.8 4 4 0.57 1/1 4 5 0.58 1/1 4 6 0.60 1/2 4 8 0.63 1/2 4 10 0.67 1/3 4 20 0.83 1/5 6 3 0.80 1/0.5 6 4 0.82 1/0.7 6 5 0.83 1/0.9 6 6 0.85 1/1 - 45 Microorganism Qa Qb SI A/B 6 8 0.88 1/1 6 10 0.92 1/2 8 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 200 1.00 ---- (4 days) 10 30 0.48 1/3 10 40 0.53 1/4 10 50 0.58 1/5 10 60 0.63 1/6 10 80 0.73 1/8 10 100 0.83 1/10 20 4 0.69 1/0.2 20 5 0.69 1/0.3 20 6 0.70 1/0.3 20 8 0.71 1/0.4 20 10 0.72 1/0.5 20 20 0.77 1/4 20 30 0.82 1/1.5 20 40 0.87 1/2 20 50 0.92 1/3 20 60 0.97 1/3 30 0 1.00

----

The ratios of MBIT/benzalkonium chloride tested ranged from 1/0.01 to 1/400. 5 Table 29 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Benzethonium chloride (Hyamine 1622) [0 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20 1.00 ---- (24 hours) 2.5 20 1.25 1/8 5 20 1.50 1/4 7.5 5 1.00 1/0.7 10 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 4 1.00 ---- (24 hours) 2 3 1.00 1/2 -46 Microorganism Qa Qb SI A/B 4 3 1.25 1/0.8 6 1 1.00 1/0.2 8 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 10 1.00 ---- (4 days) 10 4 0.73 1/0.4 10 5 0.83 1/0.5 10 6 0.93 1/0.6 30 0 1.00

----

The ratios of MBIT/benzethonium chloride tested ranged from 1/0.01 to 1/400. 5 Table 30 First Component (A) = N-methyl- 1, 2-benzisothiazolin- 3-one (MBIT) Second Component (B) = Benzyl alcohol 0 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 5000 1.00 ---- (24 hours) 2.5 5000 1.25 1/2000 5 4000 1.30 1/800 7.5 4000 1.55 1/533 10 0 1.00 --- Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 4000 1.00 ---- (48 hours) 2.5 2000 0.75 1/800 5 500 0.63 1/100 5 1000 0.75 1/200 7.5 40 0.76 1/5 7.5 50 0.76 1/7 7.5 60 0.77 1/8 7.5 80 0.77 1/11 7.5 100 0.78 1/13 7.5 200 0.80 1/27 7.5 300 0.08 1/40 7.5 400 0.85 1/53 7.5 500 0.88 1/67 -47 7.5 600 0.90 1/80 7.5 800 0.95 1/107 10 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 6000 1.00 ---- (4 days) 10 6000 1.20 1/600 20 6000 1.40 1/300 30 4000 1.27 1/133 50 0 1.00 --- The ratios of MBIT/benzyl alcohol tested ranged from 1/0.1 to 1/4000. 5 Table 31 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Benzisothiazolinone (BIT) Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20 1.00 ---- (24 hours) 2.5 10 0.75 1/4 5 5 0.75 1/1 5 6 0.80 1/1 5 8 0.90 1/2 7.5 3 0.90 1/0.4 7.5 4 0.95 1/0.5 10 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 30 1.00 ---- (48 hours) 2.5 10 0.58 1/4 2.5 20 0.92 1/8 5 4 0.63 1/0.8 5 5 0.67 1/1 5 6 0.70 1/1 5 8 0.77 1/2 5 10 0.83 1/2 7.5 2 0.82 1/0.3 7.5 3 0.85 1/0.4 7.5 4 0.88 1/0.5 7.5 5 0.92 1/0.7 -48 Microorganism Qa Qb SI A/B 7.5 6 0.95 1/0.8 10 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 300 1.00 ---- (7 days) 10 100 0.53 1/10 10 200 0.87 1/20 20 30 0.50 1/2 20 40 0.53 1/2 20 50 0.57 1/3 20 60 0.60 1/3 20 80 0.67 1/4 20 100 0.73 1/5 30 20 0.67 1/0.7 30 30 0.70 1/1 30 40 0.73 1/1 30 50 0.77 1/2 30 60 0.80 1/2 30 80 0.87 1/3 30 100 0.93 1/3 50 0 1.00

----

The ratios of MBIT/benzisothiazolinone tested ranged from 1/0.01 to 1/400. 5 Table 32 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = 2-bromo-2-nitro-propane-1, 3-diol (BNPD) Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 8 1.00 ---- (24 hours) 2.5 5 0.88 1/2 5 1 0.63 1/0.2 5 2 0.75 1/0.4 5 3 0.88 1/0.6 7.5 1 0.88 1/0.1 7.5 2 1.00 1/0.3 10 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 1000 1.00 ----- -49 Microorganism Qa Qb SI A/B (24 hours) 4 100 0.60 1/25 4 200 0.70 1/50 4 300 0.80 1/75 4 400 0.90 1/100 6 4 0.75 1/0.7 6 5 0.76 1/0.8 6 6 0.76 1/1 6 8 0.76 1/1 6 10 0.76 1/2 6 20 0.77 1/3 6 30 0.78 1/5 6 40 0.79 1/7 6 50 0.80 1/8 6 60 0.81 1/10 6 80 0.83 1/13 6 100 0.85 1/17 6 200 0.95 1/33 8 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 2000 1.00 ---- (4 days) 10 800 0.60 0/80 10 1000 0.70 1/100 20 5 0.40 1/0.3 20 6 0.40 1/0.3 20 8 0.40 1/0.4 20 10 0.41 1/0.5 20 20 0.41 1/1 20 30 0.42 1/2 20 40 0.42 1/2 20 50 0.43 1/3 20 60 0.43 1/3 20 80 0.44 1/4 20 100 0.45 1/5 20 200 0.50 1/10 20 300 0.55 1/15 20 400 0.60 1/20 20 500 0.65 1/25 20 600 0.70 1/30 20 800 0.80 1/40 -50 Microorganism Qa Qb SI A/B 20 1000 0.90 1/50 30 0 1.00

----

The ratios of MBIT/2-bromo-2-nitro-propane-1, 3-diol tested ranged from 1/0.01 to 1/400. 5 Table 33 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Butylene glycol 0 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20000 1.00 ---- (24 hours) 2.5 20000 1.33 1/8000 5 20000 1.67 1/4000 7.5 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 20000 1.00 ---- (72 hours) 8 100 0.81 1/13 8 200 0.81 1/25 8 300 0.82 1/38 8 400 0.82 1/50 8 500 0.83 1/63 8 600 0.83 1/75 8 800 0.84 1/100 8 1000 0.85 1/125 8 2000 0.90 1/250 8 3000 0.95 1/375 10 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 10000 1.00 (7 days) 10 10000 1.25 1/1000 20 10000 1.50 1/500 30 10000 1.75 1/333 40 0 1.00 --- The ratios of MBIT/butylene glycol tested ranged from 1/0.1 to 1/4000.

- 51 Table 34 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) 5 Second Component (B) = Caprylyl glycol Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 2000 1.00 ---- (24 hours) 2.5 2000 1.25 1/800 5 2000 1.50 1/400 7.5 2000 1.75 1/267 10 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 4000 1.00 ---- (24 hours) 2 2000 0.70 1/1000 2 3000 0.95 1/1500 4 2000 0.90 1/500 6 600 0.75 1/100 6 800 0.80 1/133 6 1000 0.85 1/167 8 200 0.85 1/25 8 300 0.88 1/38 8 400 0.90 1/50 8 500 0.93 1/63 8 600 0.95 1/75 10 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 1000 1.00 ---- (4 days) 10 1000 0.58 1/100 10 2000 0.83 1/200 20 1000 0.92 1/50 20 0 1.00

----

The ratios of MBIT/caprylyl glycol tested ranged from 1/0.1 to 1/4000. 10 - 52 Table 35 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Chlorphenesin 5 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 2000 1.00 ---- (24 hours) 7.5 800 0.90 1/107 15 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 2000 1.00 ---- (72 hours) 4 200 0.50 1/50 4 300 0.55 1/75 4 400 0.60 1/100 4 500 0.65 1/125 4 600 0.70 1/150 4 800 0.80 1/200 4 1000 0.90 1/250 6 80 0.64 1/13 6 100 0.65 1/17 6 200 0.70 1/33 6 300 0.75 1/50 6 400 0.80 1/67 6 500 0.85 1/83 6 600 0.90 1/100 8 40 0.82 1/5 8 50 0.83 1/6 8 60 0.83 1/8 8 80 0.84 1/10 8 100 0.85 1/13 8 200 0.90 1/25 8 300 0.95 1/38 10 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 2000 1.00 ---- (3 days) 10 2000 1.50 1/200 20 0 1.00

----

The ratios of MBIT/chlorphenesin tested ranged from 1/0.1 to 1/4000.

- 53 Table 36 First Component (A) = N-methyl-1, 2-benzisothiazolin- 3-one (MBIT) Second Component (B) = DMDM Hydantoin (DMDMH) 5 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 50 1.00 ---- (24 hours) 2.5 60 1.45 1/24 5 30 1.10 1/6 7.5 10 0.95 1/1 10 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 3000 1.00 ---- (24 hours) 2.5 3000 1.33 1/1200 5 1000 1.00 1/200 7.5 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 1000 1.00 (3 days) 10 2000 2.25 1/200 20 2000 2.50 1/100 40 0 1.00

----

The ratios of MBIT/DMDM Hydantoin tested ranged from 1/0.1 to 1/4000. No synergy was observed between MBIT and DMDM Hydantoin. 0 Table 37 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = dithio-2, 2'-bis(N-methylbenzamide) (DTBMA) [5 Microorganism Q8 Qb SI A/B E. coli8739 - M9GY 0 80 1.00 ---- (24 hours) 2.5 20 0.42 1/8 2.5 30 0.54 1/12 2.5 40 0.67 1/16 2.5 50 0.79 1/20 2.5 60 0.92 1/24 5 20 0.58 1/4 5 30 0.71 1/6 5 40 0.83 1/8 -54 Microorganism Qa Qb SI A/B 5 50 0.96 1/10 7.5 3 0.54 1/0.4 7.5 4 0.55 1/0.5 7.5 5 0.56 1/0.7 7.5 6 0.58 1/0.8 7.5 8 0.60 1/1 7.5 10 0.63 1/1 7.5 20 0.75 1/3 7.5 30 0.88 1/4 10 1 0.68 1/0.1 10 2 0.69 1/0.2 10 3 0.70 1/0.3 10 4 0.72 1/0.4 10 5 0.73 1/0.5 10 6 0.74 1/0.6 10 8 0.77 1/0.8 10 10 0.79 1/1 10 20 0.92 1/2 15 0 1.00

----

Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 2000 1.00 ---- (24 hours) 6 60 0.63 1/10 6 80 0.64 1/13 6 100 0.65 1/17 6 200 0.70 1/33 6 300 0.75 1/50 6 400 0.80 1/67 6 500 0.85 1/83 6 600 0.90 1/100 8 10 0.81 1/1 8 20 0.81 1/3 8 30 0.82 1/4 8 40 0.82 1/5 8 50 0.83 1/6 8 60 0.83 1/8 8 80 0.84 1/10 8 100 0.85 1/13 8 200 0.90 1/25 8 300 0.95 1/38 -55 Microorganism Qa Qb SI A/B 10 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 800 1.00 ---- (3 days) 10 500 0.96 1/50 30 0 1.00

----

The ratios of MBIT/ dithio-2, 2'-bis(N-methylbenzamide) tested ranged from 1/0.01 to 1/400. 5 Table 38 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Ethylenediamine tetraacetic acid (EDTA) 0 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 40000 1.00 ---- (24 hours) 5 1600 0.54 1/320 10 0 1.00

----

Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 1000 1.00 ---- (48 hours) 2.5 60 0.43 1/24 2.5 80 0.47 1/32 2.5 100 0.50 1/40 2.5 120 0.53 1/48 7.5 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 1600 1.00 ---- (3 days) 10 800 0.75 1/80 10 1000 0.88 1/100 20 200 0.63 1/10 20 400 0.75 1/20 20 600 0.88 1/30 40 0 1.00

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The ratios of MBIT/ethylenediamine tetraacetic acid tested ranged from 1/0.3 to 1/8000. 15 - 56 Table 39 First Component (A) = N-methyl-1, 2-benzisothiazolin- 3-one (MBIT) 5 Second Component (B) = Ethylparaben Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 2000 1.00 ---- (24 hours) 2.5 2000 1.25 1/800 5 2000 1.50 1/400 7.5 400 0.95 1/53 10 0 1.00

----

Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 2000 1.00 ---- (24 hours) 2.5 600 0.63 1/240 2.5 800 0.73 1/320 2.5 1000 0.83 1/400 2.5 2000 1.33 1/800 5 50 0.69 1/10 5 60 0.70 1/12 5 80 0.71 1/16 5 100 0.72 1/20 5 200 0.77 1/40 5 300 0.82 1/60 5 400 0.87 1/80 5 500 0.92 1/100 5 600 0.97 1/120 7.5 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 800 1.00 ---- (7 days) 30 200 0.85 1/7 30 300 0.98 1/10 50 0 1.00

----

The ratios of MBIT/ethylparaben tested ranged from 1/0.1 to 1/4000. 10 - 57 Table 40 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Hexamidine diisethionate 5 Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 8 1.00 - (24 hours) 2.5 8 1.17 1/3 5 8 1.33 1/2 7.5 5 1.13 1/0.7 10 3 1.04 1/0.3 15 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 10 1.00 ---- (24 hours) 2 5 0.70 1/3 2 6 0.80 1/3 4 4 0.80 1/1 4 5 0.90 1/1 8 1 0.90 1/0.1 10 0 1.00

----

The ratios of MBIT/hexamidine diisethionate tested ranged from 1/0.01 to 1/400. 0 Table 41 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Hexetidine 15 Microorganism Qa Qb SI A/B E. coli 8739 - M9GY 0 80 1.00 ---- (72 hours) 5 40 0.83 1/8 5 50 0.96 1/10 15 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 80 1.00 ---- (72 hours) 2 40 0.53 1/20 2 50 0.63 1/25 2 60 0.73 1/30 2 80 0.93 1/40 4 30 0.57 1/8 -58 Microorganism Qa Qb SI A/B 4 40 0.67 1/10 4 50 0.77 1/13 4 60 0.87 1/15 6 20 0.60 1/3 6 30 0.70 1/5 6 40 0.80 1/7 6 50 0.90 1/8 8 5 0.58 1/0.6 8 6 0.59 1/0.7 8 8 0.61 1/1 8 10 0.63 1/1 8 20 0.73 1/3 8 30 0.83 1/4 8 40 0.93 1/5 10 1 0.68 1/0.1 10 2 0.69 1/0.2 10 3 0.70 1/0.3 15 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 200 1.00 (3 days) 10 40 0.53 1/4 10 50 0.58 1/5 10 60 0.63 1/6 10 80 0.73 1/8 10 100 0.83 1/10 20 20 0.77 1/1 20 30 0.82 1/2 20 40 0.87 1/2 20 50 0.92 1/3 20 60 0.97 1/3 20 0 1.00

----

The ratios of MBIT/hexetidine tested ranged from 1/0.01 to 1/400.

- 59 Table 42 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) 5 Second Component (B) = Hexylene glycol Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20000 1.00 ---- (24 hours) 2.5 20000 1.17 1/8000 5 20000 1.33 1/4000 7.5 20000 1.50 1/2667 10 20000 1.67 1/2000 15 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 20000 1.00 ---- (24 hours) 2 20000 1.13 1/10000 4 20000 1.27 1/5000 6 20000 1.40 1/3333 8 20000 1.53 1/2500 10 20000 1.67 1/2000 15 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 20000 1.00 ---- (3 days) 10 20000 1.33 1/2000 20 20000 1.67 1/1000 30 0 1.00

----

The ratios of MBIT/hexylene glycol tested ranged from 1/0.1 to 1/10000. No synergy was observed between MBIT and hexylene glycol. 10 Table 43 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) 15 Second Component (B) = Methylparaben Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 2000 1.00 ---- (24 hours) 2.5 2000 1.25 1/800 5 2000 1.50 1/400 7.5 800 1.15 1/107 10 0 1.00

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- 60 Microorganism Qa Qb SI A/B Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 2000 1.00 ---- (48 hours) 2.5 1000 0.83 1/400 5 400 0.87 1/80 5 500 0.92 1/100 5 600 0.97 1/120 7.5 0 1.00

----

Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 800 1.00 ---- (7 days) 30 50 0.66 1/2 30 60 0.68 1/2 30 80 0.70 1/3 30 100 0.73 1/6 30 200 0.85 1/7 30 300 0.98 1/10 50 0 1.00

----

The ratios of MBIT/methylparaben tested ranged from 1/0.1 to 1/4000. 5 Table 44 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Pentylene glycol Microorganism Q. Qi SI A/B E. coli8739 - M9GY 0 20000 1.00 ---- (24 hours) 2.5 20000 1.17 1/8000 5 20000 1.33 1/4000 7.5 20000 1.50 1/2667 10 20000 1.67 1/2000 15 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 20000 1.00 ---- (24 hours) 2 20000 1.13 1/10000 4 20000 1.27 1/5000 8 20000 1.53 1/2500 10 0 1.00

----

Microorganism Qa Qb SI A/B -61 Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 20000 1.00 ---- (3 days) 10 20000 1.67 1/2000 20 20000 2.33 1/1000 30 0 1.00

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The ratios of MBIT/pentylene glycol tested ranged from 1/0.1 to 1/10000. No synergy was observed between MBIT and pentylene glycol. 5 Table 45 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Phenoxyethanol 0 Microorganism Qa Qb SI A/B E. col 8739 - M9GY 0 4000 1.00 ---- (24 hours) 2.5 4000 1.17 1/1600 5 4000 1.33 1/800 7.5 4000 1.50 1/533 10 3000 1.42 1/300 40 3000 3.42 1/75 15 0 1.00

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Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 3000 1.00 ---- (24 hours) 5 600 0.87 1/120 5 800 0.93 1/160 7.5 0 1.00

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Microorganism Q8 Qb SI A/B A. niger 16404 - PDB 0 4000 1.00 ---- (3 days) 10 4000 1.25 1/400 20 2000 1.00 1/100 40 0 1.00

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The ratios of MBIT/phenoxyethanol tested ranged from 1/0.1 to 1/4000.

- 62 Table 46 First Component (A) = N-methyl-1, 2-benzisothiazolin- 3-one (MBIT) 5 Second Component (B) = linoleamidopropyl PG-dimonium chloride phosphate (Phospholipid EFA) Microorganism Qa Qb SI A/B E coli 8739 - M9GY 0 500 1.00 ---- (48 hours) 15 40 0.83 1/3 15 50 0.85 1/3 15 60 0.87 1/4 15 80 0.91 1/5 15 100 0.95 1/7 20 0 1.00

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Microorganism Q. Qb SI A/B C. albicans 10231 - PDB 0 1000 1.00 ---- (24 hours) 2 800 0.93 1/400 4 600 0.87 1/150 6 500 0.90 1/83 15 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 2000 1.00 ---- (3 days) 10 800 0.73 1/80 10 1000 0.83 1/100 20 300 0.82 1/15 20 400 0.87 1/20 20 500 0.92 1/25 20 600 0.97 1/30 30 0 1.00

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The ratios of MBIT/linoleamidopropyl PG-dimonium chloride phosphate 10 tested ranged from 1/0.01 to 1/400.

- 63 Table 47 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = cocamidopropyl PG-dimonium chloride phosphate 5 (Phospholipid PTC) Microorganism Qa Qb SI A/B E. col 8739 - M9GY 0 100 1.00 ---- (48 hours) 2.5 60 0.73 1/24 2.5 80 0.93 1/32 10 30 0.80 1/3 10 40 0.90 1/4 15 10 0.85 1/0.7 15 20 0.95 1/1 20 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 1000 1.00 ---- (24 hours) 4 500 0.90 1/125 6 300 0.90 1/50 8 3 0.80 1/0.4 8 4 0.80 1/0.5 8 5 0.81 1/0.7 8 6 0.81 1/0.8 8 8 0.81 1/1 8 10 0.81 1/1 8 20 0.82 1/3 8 30 0.83 1/4 8 40 0.84 1/5 8 50 0.85 1/6 8 60 0.86 1/8 8 80 0.88 1/10 8 100 0.90 1/13 10 0 1.00

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Microorganism Qa Q, SI A/B A. niger 16404 - PDB 0 1000 1.00 ---- (4 days) 10 500 0.83 1/50 10 600 0.93 1/60 30 0 1.00

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The ratios of MBIT/cocamidopropyl PG-dimonium chloride phosphate tested ranged from 1/0.01 to 1/400.

- 64 Table 48 First Component (A) = N-methyl-1, 2-benzisothiazolin- 3-one (MBIT) 5 Second Component (B) = Potassium sorbate Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20000 1.00 ---- (48 hours) 10 1000 0.72 1/100 10 2000 0.77 1/200 10 3000 0.82 1/300 10 4000 0.87 1/400 10 5000 0.92 1/500 10 6000 0.97 1/600 15 0 1.00

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Microorganism Qa Qb SI A/B C a/bicans 10231 - PDB 0 400 1.00 ---- (24 hours) 2.5 200 0.83 1/80 5 40 0.77 1/8 5 50 0.79 1/10 5 60 0.82 1/12 5 80 0.87 1/16 5 100 0.92 1/20 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 10000 1.00 ---- (4 days) 10 6000 0.85 1/600 20 1000 0.60 1/50 20 2000 0.70 1/100 20 3000 0.80 1/150 20 4000 0.90 1/200 40 0 1.00

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The ratios of MBIT/potassium sorbate tested ranged from 1/0.1 to 1/4000.

- 65 Table 49 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) 5 Second Component (B) = Propylparaben Microorganism Qa Qb SI A/B E. coli 8739 - M9GY 0 4000 1.00 ---- (48 hours) 2.5 2000 0.67 1/800 2.5 3000 0.92 1/1200 5 2000 0.83 1/400 15 0 1.00

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Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 2000 1.00 ---- (24 hours) 2.5 800 0.73 1/320 2.5 1000 0.83 1/400 5 400 0.87 1/80 5 500 0.92 1/100 5 600 0.97 1/120 7.5 0 1.00

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Microorganism Q8 Qb SI A/B A. niger 16404 - PDB 0 2000 1.00 ---- (7 days) 10 800 0.60 1/80 10 1000 0.70 1/100 20 1000 0.90 1/50 30 300 0.75 1/10 30 400 0.80 1/13 30 500 0.85 1/17 30 600 0.90 1/20 50 0 1.00

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The ratios of MBIT/propylparaben tested ranged from 1/0.1 to 1/4000.

- 66 Table 50 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) 5 Second Component (B) = Quaternium-15 (active - 1-(3-chloroallyl)-3,5,7-triaza-1 azoniaadamantane chloride) Microorganism Qa Qb SI A/B E coli8739 - M9GY 0 50 1.00 ---- (24 hours) 5 20 0.90 1/400 7.5 2 0.79 1/0.3 7.5 3 0.81 1/0.4 7.5 4 0.83 1/0.5 7.5 5 0.85 1/0.7 7.5 6 0.87 1/0.8 7.5 8 0.91 1/1 7.5 10 0.95 1/1 10 0 1.00

----

Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 500 1.00 ---- (24 hours) 2.5 500 1.33 1/200 5 300 1.27 1/60 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 3000 1.00 ---- (3 days) 10 1000 0.58 1/100 10 2000 0.92 1/200 20 1000 0.83 1/50 40 0 1.00

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The ratios of MBIT/1-(3-chloroallyl)-3,5,7-triaza-1-azoniaadamantane 10 chloride tested ranged from 1/0.01 to 1/4000. Table 51 15 First Component (A) = N-methyl-1, 2-benzisothiazolin- 3-one (MBIT) Second Component (B) = Dehydroacetic acid, sodium salt (SDHA) Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 20000 1.00 ---- (24 hours) 2.5 20000 1.25 1/8000 -67 Microorganism Qa Qb SI A/B 5 20000 1.50 1/4000 7.5 20000 1.75 1/2667 10 0 1.00 Microorganism Qa Qb SI A/B C albicans 10231 - PDB 0 40 1.00 -.- (24 hours) 2 40 1.25 1/20 4 30 1.25 1/8 6 8 0.95 1/1 8 0 1.00 - Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 200 1.00 (7 days) 20 80 0.80 1/4 20 100 0.90 1/5 30 50 0.85 1/2 30 60 0.90 1/2 50 0 1.00

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The ratios of MBIT/dehydroacetic acid, sodium salt tested ranged from 1/0.01 to 1/4000. 5 Table 52 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Sodium benzoate 0 Microorganism Qa Qb SI A/B E. coli 8739 - M9GY 0 10000 1.00 ---- (24 hours) 2.5 10000 1.25 1/4000 5 10000 1.50 1/2000 7.5 10000 1.75 1/1333 10 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 2000 1.00 -- (48 hours) 2.5 800 0.65 1/320 2.5 1000 0.75 1/400 5 300 0.65 1/60 5 400 0.70 1/80 5 500 0.75 1/100 -68 Microorganism Qa Qb SI A/B 5 600 0.80 1/120 5 800 0.90 1/160 7.5 30 0.77 1/4 7.5 40 0.77 1/5 7.5 50 0.78 1/7 7.5 60 0.78 1/8 7.5 80 0.79 1/11 7.5 100 0.80 1/13 7.5 200 0.85 1/27 7.5 300 0.90 1/40 7.5 400 0.95 1/53 10 0 1.00

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Microorganism Q8 Qb SI A/B A. niger 16404 - PDB 0 10000 1.00 ---- (7 days) 10 6000 0.77 1/600 10 8000 0.97 1/800 20 6000 0.93 1/300 30 2000 0.70 1/67 30 3000 0.80 1/100 30 4000 0.90 1/133 40 2000 0.87 1/50 40 3000 0.97 1/75 60 0 1.00

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The ratios of MBIT/sodium benzoate tested ranged from 1/0.1 to 1/4000. 5 Table 53 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B)= Sodium citrate Microorganism Qa Qb SI A/B E. coli 8739 - M9GY 0 20000 1.00 ---- (24 hours) 2.5 20000 1.25 1/8000 5 20000 1.50 1/4000 7.5 20000 1.75 1/2667 10 0 1.00

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Microorganism Qa Qb SI A/B -69 Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 10000 1.00 ---- (24 hours) 2.5 3000 0.63 1/1200 2.5 4000 0.73 1/1600 2.5 5000 0.83 1/2000 2.5 6000 0.93 1/2400 5 500 0.72 1/100 5 600 0.73 1/120 5 800 0.75 1/160 5 1000 0.77 1/200 5 2000 0.87 1/400 5 3000 0.97 1/600 7.5 0 1.00

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Microorganism Qa Qb SI A/B A. niger 16404 - PDB 0 10000 1.00 ---- (7 days) 30 1000 0.70 1/33 30 2000 0.80 1/67 30 3000 0.90 1/100 50 0 1.00

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The ratios of MBIT/sodium citrate tested ranged from 1/0.1 to 1/4000. 5 Table 54 First Component (A) = N-methyl-1,2-benzisothiazolin-3-one (MBIT) Second Component (B) = Zinc pyrithione Microorganism Qa Qb SI A/B E. coli8739 - M9GY 0 2 1.00 ---- (48 hours) 2.5 0.4 0.45 1/0.2 2.5 0.5 0.50 1/0.2 2.5 0.6 0.55 1/0.2 2.5 0.8 0.65 1/0.3 2.5 1 0.75 1/0.4 5 0.3 0.65 1/0.06 5 0.4 0.70 1/0.08 5 0.5 0.75 1/0.1 5 0.6 0.80 1/0.1 5 0.8 0.90 1/0.2 7.5 0.3 0.90 1/0.04 -70 Microorganism Qa Qb SI A/B 7.5 0.4 0.95 1/0.05 10 0 1.00

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Microorganism Qa Qb SI A/B C. albicans 10231 - PDB 0 30 1.00 ---- (72 hours) 2.5 4 0.47 1/2 2.5 5 0.50 1/2 2.5 6 0.53 1/2 2.5 8 0.60 1/3 2.5 10 0.67 1/4 5 2 0.73 1/0.4 5 3 0.77 1/0.6 5 4 0.80 1/0.8 5 5 0.83 1/1 5 6 0.87 1/1 5 8 0.93 1/2 7.5 0 1.00

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A. niger 16404 - PDB 0 20 1.00 ---- (3 days) 20 5 0.75 1/3 20 6 0.80 1/0.3 20 8 0.90 1/0.4 40 0 1.00

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The ratios of MBIT/zinc pyrithione tested ranged from 1/0.001 to 1/400.

Claims (2)

1. A microbicidal composition comprising: (a) N- (n-butyl)-1, 2-benzisothiazolin- 3-one; and (b) 1,3-dimethylol-5,5-dimethyl hydantoin.

2. The composition according to claim 1, substantially as hereinbefore described with reference to any one of the Examples.